title: Isolation of binding sites to glycophorin from Mycoplasma pneumoniae membranes.
abstract: Sialoglycoproteins are major receptor sites for attachment of Mycoplasma pneumoniae to respiratory epithelium and erythrocytes (RBC). We used glycophorin, the major sialoglycoprotein of human RBC, as a ligand in affinity chromatography for the isolation of the binding sites from M. pneumoniae membranes. Membranes isolated from M. pneumoniae cells, radioiodinated by the lactoperoxidase technique, were treated with 0.5% deoxycholate. The insoluble residue, exhibiting an increased capacity to bind to RBC, was solubilized by 0.1% sodium dodecyl sulfate. The solubilized material was subjected to chromatography on a glycophorin-Sepharose column. The fraction retained on the column was eluted with 0.2% sodium dodecyl sulfate. It lacked the high-molecular-weight polypeptides and was highly enriched with two polypeptides (apparent molecular weights, 45,000 and 25,000). The eluted fraction exhibited a high capacity to bind to glycophorin-Sepharose beads and a lower capacity to bind to RBC. The binding of the eluted fraction to RBC was almost completely abolished by glycophorin, but not by its hydrophobic moiety. Binding of the fraction to glycophorin-Sepharose beads was inhibited to about the same extent by both glycophorin and its hydrophobic moiety, suggesting that components of the eluted fraction are also capable of binding to the hydrophobic moiety of glycophorin, which is apparently exposed on the beads but not on the RBC surface.
mesh_terms: D001665:Binding Sites; D002458:Cell Fractionation; D002462:Cell Membrane / Q000032:analysis; D002846:Chromatography, Affinity; D003840:Deoxycholic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D006021:Glycophorins / Q000378:metabolism* / Q000494:pharmacology; D009177:Mycoplasma pneumoniae / Q000032:analysis* / Q000378:metabolism; D012795:Sialoglycoproteins / Q000378:metabolism*; D014357:Trypsin
chemical_list: D006021:Glycophorins; D012795:Sialoglycoproteins; D003840:Deoxycholic Acid; D014357:Trypsin